The goal of this study is a better understanding of the cellular mutagenesis processes that deal with aberrant structures in template DNA and produce altered DNA sequences. Molecular activities are considered for the repair of permutational lesions and for the accommodation of non- repaired lesions that gives rise to mutations. The emphasis is on in vivo studies, exploiting the convenient genetics and accumulated background information for Escherichia coli cells and the procedures refined in this laboratory and elsewhere for efficient quantification of specific mutation consequences. Variables are considered in the molecular physiology of mutagenesis. The results will increase fundamental knowledge about how radiation alters heritable genetic treats, how certain identified parts work together and with other parts of the cell to mediate translesion mutation fixation, and how DNA repair, expression, replication and mutation interrelate at the gene. The work will be relevant to mutagenesis by various agents and to carcinogenesis and possibly processes for differentiation and diversity in the immune response. Seven focused ideas are to be investigated in three Specific Aims regarding: A. The Transcription-Repair Coupling Factor (TRCF) and Mutation Frequency Decline (MFD), B. Mutation at apurinic (AP) sites, and C. Effects on DNA replication velocity by mutagenesis proteins: A-i, TRCF may have an effect on mutation fixation apart from its effect on excision repair - knowledge about this additional role would help explain TRCF function; A-ii, The rate of MFD, when limited by saturating damage in the cell, should respond to an increased level of the limiting component and thereby identify the component and rate limiting aspect of strand- specific excision repair; A-iii, Control of MFD by amino acids in the medium may or may not be mediated by the rate of tRNA expression, which can be decided by studies of mutation in plasmids expressing tRNA from the normal or atypical promoter; B-i, Mutation fixation at AP sites requires the assembly of an appropriate structure, and components for this SOS structure distribute between different kinds of lesion in the cell - a specific limiting component can be identified by amplifying individual possibilities; B-ii, Pol II enzyme has a particular role in mutagenesis at AP sites that may be masked by the presence of other DNA lesions in the cell, e.g., by the preferred sequestering of a critical SOS component at other lesions; B-iii, The mutagenic mechanism at AP sites is highly sensitive to circumstances of the cell and even to the nature of the DNA backbone supporting the AP site, which can be studied with mutation spectra analyses; C, Certain proteins associated with UV- mutagenesis in E. coli may contribute fundamentally to the mechanism for moving replication forks along DNA (the replication velocity).